Highly pure pemetrexed Diacid and processes for the preparation thereof

ABSTRACT

Provided are highly pure pemetrexed diacid and pharmaceutically acceptable salts thereof, and processes for their preparation.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is a continuation of U.S. patent application Ser. No. 11/893,299 filed Aug. 14, 2007 and claims the benefit of priority to U.S. provisional Application Ser. No. 60/837,637, filed Aug. 15, 2006; U.S. provisional Application Ser. No. 60/860,554, filed Nov. 21, 2006; U.S. provisional Application Ser. No. 60/880,178, filed Jan. 11, 2007; U.S. provisional Application Ser. No. 60/958,213, filed Jul. 3, 2007; U.S. provisional Application Ser. No. 60/837,303, filed Aug. 14, 2006; U.S. provisional Application Ser. No. 60/860,557, filed Nov. 21, 2006; U.S. provisional Application Ser. No. 60/839,551, filed Aug. 22, 2006; U.S. provisional Application Ser. No. 60/845,031, filed Sep. 14, 2006; U.S. provisional Application Ser. No. 60/899,928, filed Feb. 6, 2007; U.S. provisional Application Ser. No. 60/936,553, filed Jun. 20, 2007; U.S. provisional Application Ser. No. 60/958,413, filed Jul. 5, 2007; U.S. provisional Application Ser. No. 60/847,291, filed Sep. 25, 2006; U.S. provisional Application Ser. No. 60/855,139, filed Oct. 30, 2006; U.S. provisional Application Ser. No. 60/880,179, filed Jan. 11, 2007; and U.S. provisional Application Ser. No. 60/958,326, filed Jul. 2, 2007, hereby incorporated by reference.

FIELD OF THE INVENTION

The invention encompasses highly pure pemetrexed diacid and pharmaceutically acceptable salts thereof, and processes for their preparation.

BACKGROUND OF THE INVENTION

Compounds known to have antifolate activity are well recognized as chemotherapeutic agents for the treatment of cancer. In particular, compounds in the folic acid family have various activities at the enzymatic level as they inhibit such enzymes as dehydrofolate reductase, folate polyglutamate synthetase, glycinamide ribonucleotide formyltransferase and thymidylate synthetase.

A series of 4-hydroxypyrrolo[2,3-d]pyrimidine-L-glutamic acid derivatives have been disclosed, for example in EP 0434426, and shown to be particularly useful antifolate drugs, among them, pemetrexed disodium heptahydrate of formula I.

Pemetrexed disodium salt heptahydrate is marketed by Eli Lilly and Company under the trade name ALIMTA® as sterile lyophilized powder for intravenous administration. This member of the folic acid family has been approved for treatment of malignant pleural mesothelioma and for second-line treatment of non small cell lung cancer. See Physicians' Desk Reference, 60th ed., pp. 1722-1728 (2006). The commercial product is reported to be lyophilized powder of heptahydrate pemetrexed disodium and mannitol.

The preparation of the commercial lyophilized or formulated lyophilized pemetrexed disodium is disclosed in U.S. Pat. No. 7,138,521. This pemetrexed disodium is prepared from N-[4-[2-(2-Amino-4,7-dihydro-4-oxo-3H-pyrrolo[2,3-d]pyrimidin-5-yl)ethyl]benzoyl]L-glutamic acid diethylester p-toluenesulfonic acid salt, which is saponified at a pH of between 2.5 to 3.5 to give N-[4-[2(2-amino-4,7-dihydro-4-oxo-1H-pyrrolo[2,3-d]pyrimidin-5-yl)ethyl]benzoyl]-L-glutamic acid (“pemetrexed diacid”), of the following Formula II:

The pemetrexed diacid is isolated as a wet cake and then combined with 2 to 3 equivalents of sodium hydroxide at a pH of between 7 and 9. The resulting pemetrexed disodium heptahydrate is then isolated from the reaction mixture by precipitation using acetone. The isolated pemetrexed disodium heptahydrate is then used to prepare the pharmaceutical composition.

Pemetrexed diacid and its preparation is believed to have been described for the first time in U.S. Pat. No. 5,344,932.

Formation and isolation of pemetrexed diacid from a mixture of water and ethanol having a pH of 2.5-3.5 is disclosed in U.S. Pat. No. 7,138,521. A similar isolation is disclosed in C. J. Barnett, et al., “A Practical Synthesis of Multitargeted Antifolate LY231514,” Organic Process Research & Development, 3(3): 184-188 (1999).

Formation and isolation of pemetrexed diacid from an aqueous solution having a pH of 5 is disclosed in U.S. Pat. No. 5,416,211.

Formation and isolation of pemetrexed diacid from an aqueous solution having a pH of 2.8-3.1 is disclosed in U.S. Pat. No. 6,262,262.

Like any synthetic compound, pemetrexed diacid can contain extraneous compounds or impurities that can come from many sources. They can be unreacted starting materials, by-products of the reaction, products of side reactions, or degradation products. Impurities in pemetrexed diacid as in its active pharmaceutical ingredient (API), pemetrexed disodium, are undesirable and might even be harmful to a patient being treated with a dosage form containing the API.

The purity of the API produced in the commercial manufacturing process is clearly a necessary condition for commercialization. Impurities introduced during commercial manufacturing processes must be limited to very small amounts, and are preferably substantially absent. For example, the International Conference on Harmonisation of Technical Requirements for Registration of Pharmaceuticals for Human Use (“ICH”) Q7A guidance (dated Nov. 10, 2000) for API manufacturers requires that process impurities be maintained below set limits by specifying the quality of raw materials, controlling process parameters, such as temperature, pressure, time, and stoichiometric ratios, and including purification steps, such as crystallization, distillation, and liquid-liquid extraction, in the manufacturing process.

The product mixture of a chemical reaction is rarely a single compound with sufficient purity to comply with pharmaceutical standards. Side products and by-products of the reaction and adjunct reagents used in the reaction will, in most cases, also be present in the product mixture. At certain stages during processing of an pemetrexed diacid or during the processing of an API, such as pemetrexed disodium, it must be analyzed for purity, typically, by HPLC or TLC analysis, to determine if it is suitable for continued processing and, ultimately, for use in a pharmaceutical product. The pemetrexed diacid or API need not be absolutely pure, as absolute purity is a theoretical ideal that is typically unattainable. Rather, purity standards are set with the intention of ensuring that an API is as free of impurities as possible, and thus, is as safe as possible for clinical use. As discussed above, in the United States, the Food and Drug Administration guidelines recommend that the amounts of some impurities be limited to less than 0.1 percent.

Thus, providing highly pure pemetrexed diacid and means for the preparation thereof is desirable.

SUMMARY OF THE INVENTION

In one embodiment, the invention encompasses pemetrexed diacid of the following formula,

having a purity of at least about 99%.

In another embodiment, the invention encompasses a process for preparing pemetrexed diacid having a purity of at least about 99% comprising: a) measuring the purity of the pemetrexed diacid in at least one batch of pemetrexed diacid; b) selecting a batch of pemetrexed diacid having a purity of at least about 98%; and c) preparing pemetrexed diacid having a purity of at least about 99% from the selected batch.

In another embodiment, the invention encompasses a process for preparing a pharmaceutically acceptable salt of pemetrexed comprising preparing pemetrexed diacid having a purity of at least 99% by the above-described process, and converting it to a pharmaceutically acceptable salt of pemetrexed diacid. Preferably, the pharmaceutically acceptable salt of pemetrexed diacid is pemetrexed disodium.

In yet another embodiment, the invention encompasses a process for preparing a formulated pharmaceutically acceptable salt of pemetrexed diacid comprising preparing pemetrexed diacid having a purity of at least about 99% by a process comprising crystallizing pemetrexed diacid from a mixture comprising at least one solvent selected from the group consisting of amides, pyrrolidinones, and sulfoxides, at least one anti-solvent selected from the group consisting of alcohols, esters, ketones, water, halogenated hydrocarbons, aromatic hydrocarbons, and ethers, and optionally at least one co-solvent selected from the group consisting of nitriles, wherein the starting pemetrexed diacid has a purity of at least 98%; and formulating the obtained pemetrexed diacid into a formulation comprising a pharmaceutically acceptable salt of pemetrexed diacid. Preferably, the pharmaceutically acceptable salt of pemetrexed diacid is pemetrexed disodium.

DETAILED DESCRIPTION OF THE INVENTION

The invention is directed to highly pure pemetrexed diacid and to processes for obtaining highly pure pemetrexed diacid. The benefit of purifying pemetrexed diacid is to provide a pure starting material for the preparation of the API, which is the disodium salt, or any pharmaceutically acceptable salt, thus avoiding the purification at the stage of the final product. Purification at the stage of the final pemetrexed disodium is challenging because pemetrexed disodium is unstable to high pH values and temperatures for extended periods of time.

As used herein, unless otherwise defined, the term “high purity” or “highly pure,” when referring to pemetrexed diacid or a pharmaceutically acceptable salt of pemetrexed, means a purity of at least about 99%. Preferably, the pemetrexed diacid or pharmaceutically acceptable salt of pemetrexed has a purity of at least about 99.4%, more preferably at least about 99.7%, even more preferably at least about 99.8%, and most preferably at least about 99.9%.

The invention provides pemetrexed diacid of the following formula,

having a purity of at least about 99%. Preferably, the pemetrexed diacid has a purity of at least about 99.4%, more preferably at least about 99.7%, even more preferably at least about 99.8%, and most preferably at least about 99.9%.

The purity of the pemetrexed diacid can be measured by area % units. Preferably, the purity is measured by area % units by HPLC. An exemplary HPLC method comprises: (a) combining a pemetrexed diacid sample with a solution of NaH₂PO₄ to obtain a solution; (b) injecting the solution into a 150×4.6 mm×3 μm Phenyl C18 or Octadecyl (C18) reversed phase silica gel (or similar) column; (c) eluting the sample from the column using a mixture of an aqueous solution of trifluoroacetic acid (referred to as eluent A) and acetonitrile (referred to as eluent B); and (d) measuring the pemetrexed diacid content in the relevant sample with a UV detector.

Preferably, the NaH₂PO₄ solution has a pH of about 7.5.

The eluent used may be a mixture of eluent A and eluent B, wherein the ratio of components in the mixture varies over time, in other words, a gradient eluent. Preferably, at time 0 minutes, the eluent contains 85% of eluent A and 15% of eluent B; at 2 minutes, the eluent contains 85% of eluent A and 15% of eluent B; at 30 minutes, the eluent contains 40% of eluent A and 60% of eluent B; at 35 minutes, the eluent contains 40% of eluent A and 60% of eluent B; and at 36 minutes, the eluent contains 85% of eluent A and 15% of eluent B.

Preferably, the UV detector is set at a wavelength of about 254 nm.

In another embodiment, the invention encompasses a process for preparing pemetrexed diacid having a purity of at least about 99%. The process comprises: a) measuring the purity of the pemetrexed diacid in at least one batch of pemetrexed diacid; b) selecting a batch of pemetrexed diacid having a purity of at least about 98%; and c) preparing pemetrexed diacid having a purity of at least about 99% from the selected batch.

Preferably, the pemetrexed diacid having a purity of at least about 99% is prepared from the batch of pemetrexed diacid having a purity of at least about 98% by a process comprising crystallizing pemetrexed diacid from a mixture comprising at least one solvent selected from the group consisting of amides, pyrrolidinones, and sulfoxides, and at least one anti-solvent selected from the group consisting of alcohols, esters, ketones, water, halogenated hydrocarbons, aromatic hydrocarbons, and ethers; wherein the starting pemetrexed diacid has a purity of at least about 98%.

Preferably, the crystallized pemetrexed diacid has a purity of at least about 99.4%, more preferably at least about 99.7%, even more preferably at least about 99.8%, and most preferably at least about 99.9%.

The ratio of the solvent to the anti-solvent can be about 1:3 to about 1:6 vol:vol. Alternatively, the ratio of the solvent to the anti-solvent can be about 3:1 to about 1:3 vol:vol.

As to the solvent, preferably, the amide is a C₂₋₄ amide. Preferably, the C₂₋₄ amide is dimethylformamide (“DMF”) or dimethylacetamide (“DMA”). Preferably, the pyrrolidinone is C₄₋₆ pyrrolidinone. Preferably, the C₄₋₆ pyrrolidinone is N-methyl pyrrolidinone (“NMP”). A preferred sulfoxide is a C₂₋₄ sulfoxide. Preferably, the C₂₋₄ sulfoxide is dimethylsulfoxide (“DMSO”). More preferably, the solvent is either DMSO or DMF.

As to the anti-solvent, preferably, the alcohol is a C₁₋₄ alcohol. A preferred C₁₋₄ alcohol is methanol, ethanol, propanol, isopropanol, or butanol. Preferably, the ester is a C₃₋₆ ester. A preferred C₃₋₆ ester is ethyl acetate, methyl acetate, or isobutyl acetate. Preferably, the ketone is a C₃₋₆ ketone. A preferred C₃₋₆ ketone is a C₃₋₄ ketone and more preferably acetone. Preferably, the halogenated hydrocarbon is a C₁₋₃ halogenated hydrocarbon. A preferred C₁₋₃ halogenated hydrocarbon is a C₁₋₂ halogenated, preferably chlorinated, hydrocarbon and more preferably dichloromethane (“DCM”). Preferably, the aromatic hydrocarbon is a C₆₋₉ aromatic hydrocarbon. A preferred C₆₋₉ aromatic hydrocarbon is a C₆₋₇ aromatic hydrocarbon and more preferably toluene. Preferably, the ether is a C₄₋₈ ether. A preferred C₄₋₈ ether is a C₄₋₆ ether, and more preferably tetrahydrofuran (“THF”). Most preferably, the anti-solvent is ethanol, methanol or a mixture thereof.

Preferred combinations of solvent and anti-solvent are: DMSO and ethanol, DMSO and a mixture of water and methanol, and DMF and a mixture of water and methanol. Preferably, the mixture of water and methanol contains methanol and water in a volume ratio of about 1:3 to about 3:1, respectively. More preferably, the mixture of water and methanol contains methanol and water in a volume ratio of about 1:2 to about 2:1, and most preferably about 1:1, respectively.

The solvent can optionally comprise at least one co-solvent. The co-solvent is typically a solvent that, when used in mixture with the solvent, produces a mixture in which pemetrexed diacid has increased solubility relative to the solubility of pemetrexed diacid in the solvent alone. Suitable co-solvents include nitriles. Preferably, the nitrile is a C₂₋₄ nitrile. A preferred C₂₋₄ nitrile is acetonitrile (“ACN”).

Typically, the crystallization is done by a process comprising dissolving the pemetrexed diacid having a purity of at least about 98% in the solvent to form a solution; admixing the solution with the anti-solvent to form a precipitate of pemetrexed diacid; and recovering pemetrexed diacid having a purity of at least about 99% from the precipitate.

When the pemetrexed diacid having a purity of at least about 98% and the solvent are combined, typically a mixture is obtained. The mixture may be heated to obtain the solution. Preferably, the mixture is heated to a temperature of about 25° C. to about 75° C., and more preferably to a temperature of about 35° C. to about 55° C.

Preferably, the anti-solvent is added to the solution. Preferably, the anti-solvent is added drop-wise to the solution. Preferably, the drop-wise addition is done over a period of about 1 hour to about 5 hours, more preferably, for about 1 hour to about 2.5 hours. Preferably, the anti solvent is added to the solution at a temperature of about 30° C. to about 65° C., and more preferably at about 40° C. to about 50° C.

Typically, the addition of the anti-solvent to the solution results in the formation of a suspension having a precipitate of pemetrexed diacid. Preferably, the suspension is further maintained in order to increase the yield of the precipitated pemetrexed diacid. The suspension is preferably maintained at about 30° C. to about 65° C. in order to increase the selectivity of the crystallization, and, thus, the purity of the crystallized pemetrexed diacid. Preferably, the suspension is maintained for about 20 minutes to about three hours and more preferably for about 30 minutes to about 1.5 hours. Preferably, the suspension is maintained while being stirred.

The yield of the precipitated pemetrexed diacid may be increased by cooling the suspension, and maintaining it for a period of time. Preferably, the maintained suspension is cooled to a temperature of about 30° C. to about 0° C. and more preferably to about 25° C. to about 10° C. Preferably, cooling is done over a period of about 1 to about 6 hours, and more preferably for about 2 to about 4 hours. Preferably, the cooled suspension is maintained at a temperature of about 30° C. to about 0° C. Preferably, the cooled suspension is maintained for about 0.5 hour to about 6 hours, and more preferably for about 2 to about 4 hours, prior to recovering the precipitated pemetrexed diacid.

The precipitated pemetrexed diacid may be recovered from the suspension by any method known to a skilled artisan. Preferably, the recovery is done by filtering the precipitate from the cooled maintained suspension, washing the precipitate, and drying.

Optionally, the process may further comprise filtering the precipitate from the cooled suspension and slurrying the filtered precipitate in the anti-solvent, prior to recovering the purified product. The slurrying typically decreases the level of residual solvents in the precipitate. Typically, the filtered precipitate is washed with an anti-solvent prior to slurrying it. The preferred anti-solvent for washing the precipitate is water or a mixture of water and methanol. Optionally, the washed precipitate may be dried prior to slurrying it. The drying may be done at a temperature of about 40° C. under vacuum. Preferably, slurrying is done at a temperature of about 5° C. to about 65° C. and more preferably at about 35° C. to about 50° C.

Preferably, the anti-solvent used for slurrying the product is water, ethanol, methanol or a mixture thereof. Preferably, slurrying is done for about 0.5 hour to about 4 hours and more preferably for about 1 hour to about 2 hours. Usually, the heated slurry is cooled prior to recovering the product. Preferably, the slurry is cooled to a temperature of about 30° C. to about 5° C., and more preferably about 25° C. to about 10° C., prior to the recovery process. The recovery may be done by any method known to a skilled artisan. Preferably, the recovery includes filtering the cooled slurry, and drying the filtered precipitate.

The pemetrexed diacid having a purity of at least about 99% obtained by the above-described process may be converted into the API pemetrexed disodium or any other pharmaceutically acceptable salt of pemetrexed diacid.

Preferably, the obtained pharmaceutically acceptable salt has a purity of at least about 99%, more preferably at least about 99.4%, even more preferably at least about 99.7%, even more preferably at least about 99.8%, and most preferably at least about 99.9%. The purity of the obtained pharmaceutically acceptable salt can be measured by area % units. Preferably, the purity is measured by area % units by HPLC.

The pemetrexed diacid may be converted to pemetrexed disodium, for example, according to the process disclosed in U.S. publication No. 2003/0216416, hereby incorporated by reference.

Applicants herein demonstrate that the purity of the pemetrexed diacid obtained by the processes of the present invention is retained when the pemetrexed diacid is converted into the API pemetrexed disodium, as exemplified in example 6. Thus, it is advantageous to use a highly pure pemetrexed diacid to prepare the API pemetrexed disodium, or any other pharmaceutically acceptable salt of pemetrexed.

The pemetrexed diacid having a purity of at least about 99% obtained by the above-described process may be formulated into a formulation comprising the API pemetrexed disodium or any other pharmaceutically acceptable salt of pemetrexed diacid.

Preferably, the pharmaceutically acceptable salt in the formulation, has a purity of at least about 99%, more preferably at least about 99.4%, even more preferably at least about 99.7, even more preferably at least about 99.8%, and most preferably at least about 99.9%. Preferably, the pharmaceutically acceptable salt in the formulation is lyophilized disodium salt of pemetrexed diacid.

Applicants herein also demonstrate that the purity of the pemetrexed diacid obtained by the processes of the present invention is retained in the formulated API pemetrexed disodium, which is the lyophilized disodium salt of pemetrexed, as exemplified in example 6. Thus, it is advantageous to use a highly pure pemetrexed diacid to prepare the formulated API pemetrexed disodium, or any other formulated pharmaceutically acceptable salt of pemetrexed.

The formulated pemetrexed disodium may be prepared, for example, according to the process disclosed in co-pending U.S. application Ser. No. 11/___,___, filed Aug. 14, 2007 [attorney docket no. 13150/48305, entitled “Processes for the Preparation of Lyophilized Pharmaceutically Acceptable Salts of Pemetrexed Diacid,” as well as U.S. application Ser. No. 60/847,291, filed Sep. 25, 2006; U.S. application Ser. No. 60/855,139, filed Oct. 30, 2006; U.S. application Ser. No. 60/880,179, filed Jan. 11, 2007; and U.S. application Ser. No. 60/958,326 filed Jul. 2, 2007, herein incorporated by reference.

The pemetrexed diacid having a purity of at least about 99% or salt thereof can be converted into a lyophilized pharmaceutically acceptable salt of pemetrexed by a process comprising: combining the pemetrexed diacid having a purity of at least about 99% or a salt thereof, an agent capable of forming a pharmaceutically acceptable salt of pemetrexed, and a solvent comprising water or a mixture of water and a solvent suitable for lyophilization to obtain a mixture comprising a pharmaceutically acceptable salt of pemetrexed; and removing the solvent by lyophilization to obtain a lyophilized pharmaceutically acceptable salt of pemetrexed; wherein the pharmaceutically acceptable salt of pemetrexed is not isolated prior to the lyophilization process; and the pharmaceutically acceptable salt of pemetrexed is a di-base-addition salt of pemetrexed, with the proviso that the di-base- addition salt of pemetrexed is not the same as the starting pemetrexed diacid salt.

As used herein, unless otherwise, defined, an “agent capable of forming a pharmaceutically acceptable salt of pemetrexed” refers to an agent that is capable of forming a base-addition salt of pemetrexed. Base-addition salts include, but are not limited to, alkali or alkaline earth metal salts, such as sodium, potassium, lithium, and calcium salts.

Preferably, the agent capable of forming a pharmaceutically acceptable salt of pemetrexed is an alkali (preferably sodium) or alkaline earth metal hydroxide, carbonate, phosphate, or sulfate. More preferably, the agent capable of forming a pharmaceutically acceptable salt of pemetrexed is an alkali or alkaline earth metal hydroxide, and most preferably sodium hydroxide.

The starting pemetrexed diacid salt can be an acid-addition salt or a base-addition salt. The salt can be a di salt or a mono salt. Preferably the starting pemetrexed diacid salt is a mono salt.

As used herein, unless otherwise defined, the term “mono,” with reference to a pemetrexed diacid salt, refers to a salt having only one counter cation or anion. When the salt is basic the counter cation is originated from a base. When the salt is acidic, the counter anion is originated from an acid.

Examples of mono base-addition salts of pemetrexed diacid include, but are not limited to, alkali or alkaline earth metal salts, such as sodium, potassium, lithium, and calcium salts. Preferably, the mono base-addition salt is a sodium salt.

Examples of mono acid-addition salts of pemetrexed diacid include, but are not limited to, HBr, HCl, H₂SO₄, H₃PO₄, and CH₃SO₃H salts. Preferably, the mono acid-addition salt is a HCl salt.

As used herein, unless otherwise defined, the term “di,” with reference to a pemetrexed diacid salt, refers to a salt having two counter cations. Preferably, the di-salt is a di-base-addition salt where these cations are originated from at least one base.

Preferably, the di-salt of pemetrexed diacid is selected from a group consisting of sodium, potassium, lithium, and calcium. More preferably, the di-salt is a disodium salt.

In a preferred embodiment, initially, the pemetrexed diacid having a purity of at least about 99% or salt thereof is combined with the solvent to obtain a first mixture. Preferably, the solvent is water. When a mixture of water and a solvent suitable for lyophilization is used, the solvent suitable for lyophilization may include, but is not limited to, tert-butanol, dimethylsulfoxide, or 1,4-dioxane. Preferably, the solvent suitable for lyophilization is tert-butanol.

The first mixture is then admixed with an alkali or alkaline earth metal hydroxide to obtain the mixture comprising the pharmaceutically acceptable salt of pemetrexed, and optionally a salt generated from a reaction between the alkali or alkaline earth metal hydroxide and a mono acid-addition salt of pemetrexed diacid, depending on the starting material. When the starting material is a mono acid-addition salt of pemetrexed diacid, the mixture comprising the pharmaceutically acceptable salt of pemetrexed also includes a salt, which is also a pharmaceutically acceptable salt, generated from a reaction between the alkali or alkaline earth metal hydroxide and the mono acid-addition salt of pemetrexed diacid.

Preferably, the alkali or alkaline earth metal hydroxide is NaOH, KOH, LiOH, or Ca(OH)₂, and more preferably, NaOH. Accordingly, the salt generated from the alkali or alkaline earth metal hydroxide and the mono acid-addition salt of pemetrexed diacid is preferably, NaBr, NaCl, KBr, KCl, LiBr, LiCl, CaBr₂, or CaCl₂.

Preferably, the alkali or alkaline earth metal hydroxide is added to the first mixture. Preferably, the mixture is a solution.

Typically, to obtain the pharmaceutically acceptable salt of pemetrexed from the pemetrexed diacid or salt thereof, at least about 1 mole equivalent of alkali or alkaline earth metal hydroxide per mole equivalent of the starting pemetrexed diacid or salt thereof is used. Preferably, the alkali hydroxide is used in an amount of about 1 to about 3 mole equivalents per mole equivalent of the starting pemetrexed diacid or salt thereof.

The alkali hydroxide may be in solution or solid form. Preferably, the alkali or alkaline earth metal hydroxide is in the form of an aqueous solution. Preferably, the aqueous solution of the alkali or alkaline earth metal hydroxide is a standard solution. As used herein, unless otherwise defined, the term “standard solution” refers to a solution having a known concentration, where the concentration is determined by various methods known to a skilled artisan, such as titration with acids. Preferably, the standard solution of the alkali or alkaline earth metal hydroxide has a concentration of about 0.5 M to about 4 M, more preferably, of about 2 M.

Preferably, admixing the first mixture and the alkali or alkaline earth metal hydroxide solution is performed at a temperature of about 1° C. to about 100° C., more preferably at about 10° C. to about 60° C., and most preferably at about 15° C. to about 40° C.

As a skilled artisan will appreciate, complete dissolution or a stable pH value of about 7.0 to 7.5, or, more preferably, 7.1 to 7.2, is an indication that the reaction has completed.

The process for preparing lyophilized pharmaceutically acceptable salt of pemetrexed may further comprise a process of adjusting the pH to obtain a pH of about 7.0 to about 10.0, preferably, of about 7.0 to about 9.0, more preferably, of about 7.0 to about 8.0, and most preferably, of about 7.0 to about 7.5, prior to lyophilizing the solution comprising the pharmaceutically acceptable salt of pemetrexed. The pH can be adjusted by admixing the solution comprising the pharmaceutically acceptable salt of pemetrexed, and optionally the additional pharmaceutically acceptable salt described before, with alkali hydroxide or with any one of the starting materials, i.e., pemetrexed diacid, and its salts, depending on the pH of the solution. Typically, the pH measurement is done by using a pH-meter.

Optionally, a dispersing agent may be added to the mixture before removing the solvent. Preferably, the dispersing agent is a sugar such as lactose, fructose or mannitol. Preferably, the sugar is mannitol. Preferably, the dispersing agent is present in an equimolar amount in weight relative to the amount of the pharmaceutically acceptable salt of pemetrexed.

Having described the invention with reference to certain preferred embodiments, other embodiments will become apparent to one skilled in the art from consideration of the specification. The invention is further defined by reference to the following examples describing in detail the preparation and analysis of pemetrexed diacid and pemetrexed disodium. It will be apparent to those skilled in the art that many modifications, both to materials and methods, may be practiced without departing from the scope of the invention.

EXAMPLES

Instrumentation:

HPLC method for monitoring the purity of pemetrexed diacid:

Column & Packing: GEMINI C18; 110Å; 3 μm; 150X4.6 mm (PHENOMENEX P.N. 00F-4439-EO) or equivalent Eluent A: 3 mL of TFA in 1000 mL of water Eluent B: Acetonitrile Gradient Time (min) % Eluent A % Eluent B  0 85 15  2 85 15 30 40 60 35 40 60 36 85 15 Run time 35 minutes Equilibrium time:  5 minutes Flow Rate: 1.0 mL/mins. Detector: UV at 254 nm Column 25° C. temperature: Injection 5 μl Diluent NaH₂PO₄ 4 g/L in water at pH = 7.5 Detection Limit: 0.003% area. Mobile phase composition and flow rate may be varied in order to achieve the required system suitability.

Example 1 Purification of N-[4-[2-(2-Amino-4,7-dihydro-4-oxo-1H-pyrrolo[2,3-d]pyrimidin-5-yl)ethyl]benzoyl]-L-glutamic Acid

A flask was charged with N-[4-[2-(2-amino-4,7-dihydro-4-oxo-1H-pyrrolo[2,3-d]pyrimidin-5-yl)ethyl]benzoyl]-L-glutamic acid (98.45% HPLC purity,) (2.39 g) and dimethylsulfoxide (7 ml), heated to 45° C. and stirred until complete dissolution. Absolute ethanol (14 ml) was then added drop wise in about 80 min., the obtained suspension was stirred for 20 min. at 45° C. and then cooled to 24° C. in about 2 hours. After stirring at 24° C. for 2 hours, the suspension was filtered and the solid was washed with absolute ethanol (7 ml). The wet solid was dried at 40° C. under vacuum (18 mbar) for about 16 hours affording the title compound as a light-blue solid (2.27 g) in 99.07% purity.

Example 2 Purification of N-[4-[2-(2-Amino-4,7-dihydro-4-oxo-1H-pyrrolo[2,3-d]pyrimidin-5-yl)ethyl]benzoyl]-L-glutamic Acid

A flask was charged with N-[4-[2-(2-amino-4,7-dihydro-4-oxo-1H-pyrrolo[2,3-d]pyrimidin-5-yl)ethyl]benzoyl]-L-glutamic acid (99.07% HPLC purity) (2.25 g) and dimethylsulfoxide (6.75 ml), heated to 45° C. and stirred until complete dissolution. Absolute ethanol (20.25 ml) was then added drop wise in about 2 hours, the obtained suspension was stirred for 1 hour at 45° C. and then cooled to 30° C. in about 2 hours. The suspension was filtered and the solid was washed with water (20.25 ml). The wet solid was slurried at 45° C. in absolute ethanol (20.25 ml) for one hour, cooled to 30° C. and filtered. After drying at 40° C. under vacuum (18 mbar) for about 16 hours, the title compound was obtained in 99.81% purity (HPLC).

Example 3 Purification of N-[4-[2-(2-Amino-4,7-dihydro-4-oxo-1H-pyrrolo[2,3-d]pyrimidin-5-yl)ethyl]benzoyl]-L-glutamic Acid

A flask was charged with N-[4-[2-(2-amino-4,7-dihydro-4-oxo-1H-pyrrolo[2,3-d]pyrimidin-5-yl)ethyl]benzoyl]-L-glutamic acid (98.22% HPLC purity) (10.0 g) and dimethylsulfoxide (25 ml), heated to 65° C. and stirred until complete dissolution. Methanol (37.5 ml) and water (37.5 mL) was then added drop wise in about 2.5 hours, the obtained suspension was stirred for 1 hour at 65° C. and then was cooled to 22° C. in about 2.5 hours. The suspension was filtered and the solid was washed with a mixture of water (50 ml) and methanol (50 mL). After drying at 40° C. under vacuum (18 mbar) for about 16 hours, the title compound was obtained (7.62 g). The product was dried in oven and slurried in 60 mL of water, the suspension was heated at 50° C. for 1 h and then slowly cooled to ambient and filtered. The resulting solid was dried in the oven at 80° C. under vacuum (18 mbar) for about 16 hours 99.47% purity (HPLC).

Example 4 Purification of N-[4-[2-(2-Amino-4,7-dihydro-4-oxo-1H-pyrrolo[2,3-d]pyrimidin-5-yl)ethyl]benzoyl]-L-glutamic Acid

A flask was charged with crude N-[4-[2-(2-amino-4,7-dihydro-4-oxo-1H-pyrrolo[2,3-d]pyrimidin-5-yl)ethyl]benzoyl]-L-glutamic acid (99.40% HPLC) (2.40 g) and dimethylformamide (4.8 ml), heated to 50° C. and stirred until complete dissolution. A mixture of water (7.2 ml) and methanol (7.2 ml) was then added dropwise in about 75 min., the obtained suspension was stirred for 30 min. at 50° C. and then cooled to 27° C. in about 4 hours. The suspension was filtered and the solid was washed with water (20 ml).

After drying at 40° C. under vacuum (18 mbar) for about 16 hours, the title compound was obtained (2.25 g). The product was slurried in water at 50° C. for one hour, then cooled to 26° C. in about 2 hours. The suspension was filtered. The wet solid was washed with water and dried at 40° C. under vacuum (18 mbar) for about 16 hours affording the title compound as a white solid in 99.44% purity (HPLC).

Example 5 Lyophilized N-[4-[2-(2-amino-4,7-dihydro-4-oxo-1H-pyrrolo[2,3-d]pyrimidin-5-yl)ethyl]benzoyl]-L-glutamic Acid Disodium Salt

5 grams of N-[4-[2-(2-amino-4,7-dihydro-4-oxo-1H-pyrrolo[2,3-d]pyrimidin-5-yl)ethyl]benzoyl]-L-glutamic acid (purity 99.6% by HPLC) was dissolved in 1 L of distilled water and 11.698 ml of a 2.0 M solution of sodium hydroxide was added to the solution. Mannitol (10 g) was then added to the solution and dissolved. The solution was then filtered through a bacterial filter and dried in a freeze-drier to afford the title compound as a white solid (15.5 g purity 99.6% area by HPLC).

Example 6 Preparation of N-[4-[2-(2-Amino-4,7-dihydro-4-oxo-1H-pyrrolo[2,3-d]pyrimidin-5-yl)ethyl]benzoyl]-L-glutamic Acid Disodium Salt

A flask was charged with crude N-[4-[2-(2-amino-4,7-dihydro-4-oxo-1H-pyrrolo[2,3-d]pyrimidin-5-yl)ethyl]benzoyl]-L-glutamic acid (99.50% HPLC purity) (4 g), water (28 ml) and NaOH (3 equivalents) at 25° C. Complete dissolution was obtained. The pH was corrected to 10.0 with HCl 1M and acetone (120 mL) was added drop wise over a period of 50 min. The obtained suspension was cooled to 2° C. in one hour, stirred at 2° C. for 10 hours and filtered. The solid was washed with acetone (30 mL) and dried at 40° C. under vacuum (18 mbar) for about 16 hours affording the title compound in 99.53% purity by HPLC.

Example 7 Purification of N-[4-[2-(2-Amino-4,7-dihydro-4-oxo-1H-pyrrolo[2,3-d]pyrimidin-5-yl)ethyl]benzoyl]-L-glutamic Acid

A flask was charged with N-[4-[2-(2-amino-4,7-dihydro-4-oxo-1H-pyrrolo[2,3-d]pyrimidin-5-yl)ethyl]benzoyl]-L-glutamic acid (98.22% HPLC purity) (10.0 g) and dimethylsulfoxide (25 ml), heated to 65° C. and stirred until complete dissolution. Methanol (75 ml) and water (75 mL) was then added drop wise over a period of about 2.5 hours, the obtained suspension was stirred for 1 hour at 65° C. and then was cooled to 22° C. over a period of about 2.5 hours. The suspension was filtered and the solid was washed with a mixture of water (50 ml) and methanol (50 mL). After drying at 40° C. under vacuum (18 mbar) for about 16 hours, the title compound was obtained (7.62 g).

The product was dried in oven and slurried in 60 mL of water, the suspension was heated at 50° C. for 1 h and then slowly cooled to ambient and filtered. The resulting solid was dried in the oven at 80° C. under vacuum (18 mbar) for about 16 hours 99.03% purity (HPLC).

Example 8 Purification of N-[4-[2-(2-Amino-4,7-dihydro-4-oxo-1H-pyrrolo[2,3-d]pyrimidin-5-yl)ethyl]benzoyl]-L-glutamic Acid

A flask was charged with N-[4-[2-(2-amino-4,7-dihydro-4-oxo-1H-pyrrolo[2,3-d]pyrimidin-5-yl)ethyl]benzoyl]-L-glutamic acid (99.07% HPLC purity) (2.25 g) and dimethylsulfoxide (7 ml), heated to 45° C. and stirred until complete dissolution. Absolute ethanol (5 ml) was then added drop wise over a period of about 2 hours, the obtained suspension was stirred for 1 hour at 45° C. and then cooled to 30° C. over a period of about 2 hours. The suspension was filtered and the solid was washed with water (20.25 ml). The wet solid was slurried at 45° C. in absolute ethanol (20.25 ml) for one hour, cooled to 30° C. and filtered. After drying at 40° C. under vacuum (18 mbar) for about 16 hours, the title compound was obtained in 99.89% purity (HPLC).

Example 9 Purification of N-[4-[2-(2-Amino-4,7-dihydro-4-oxo-1H-pyrrolo[2,3-d]pyrimidin-5-yl)ethyl]benzoyl]-L-glutamic Acid

A flask was charged with N-[4-[2-(2-amino-4,7-dihydro-4-oxo-1H-pyrrolo[2,3-d]pyrimidin-5-yl)ethyl]benzoyl]-L-glutamic acid (99.07% HPLC purity) (2.25 g) and dimethylsulfoxide (7 ml), heated to 45° C. and stirred until complete dissolution. Absolute ethanol (5 ml) was then added drop wise over a period of about 2 hours, the obtained suspension was stirred for 1 hour at 45° C. and then cooled to 10° C. over a period of about 2 hours. The suspension was filtered and the solid was washed with water (20.25 ml). The wet solid was slurried at 45° C. in absolute ethanol (20.25 ml) for one hour, cooled to 30° C. and filtered. After drying at 40° C. under vacuum (18 mbar) for about 16 hours, the title compound was obtained in 99.75% purity (HPLC).

Example 10 Purification of N-[4-[2-(2-Amino-4,7-dihydro-4-oxo-1H-pyrrolo[2,3-d]pyrimidin-5-yl)ethyl]benzoyl]-L-glutamic Acid

A flask was charged with crude N-[4-[2-(2-amino-4,7-dihydro-4-oxo-1H-pyrrolo[2,3-d]pyrimidin-5-yl)ethyl]benzoyl]-L-glutamic acid (99.40% HPLC purity) (2.40 g) and dimethylformamide (4.8 ml), heated to 50° C. and stirred until complete dissolution. A mixture of water (4.8 ml) and methanol (4.8 ml) was then added drop wise over a period of about 75 min., the obtained suspension was stirred for 30 minutes at 50° C. and then cooled to 24° C. over a period of about 4 hours. The suspension was filtered and the solid was washed with water (20 ml).

After drying at 40° C. under vacuum (18 mbar) for about 16 hours, the title compound was obtained (1.87 g). The product was slurried in water at 50° C. for one hour, then cooled to 24° C. in about 2 hours. The suspension was filtered. The wet solid was washed with water and dried at 40° C. under vacuum (18 mbar) for about 16 hours affording the title compound as a white solid in 99.71% purity (HPLC).

Example 11 Preparation of N-[4-[2-(2-Amino-4,7-dihydro-4-oxo-1H-pyrrolo[2,3-d]pyrimidin-5-yl)ethyl]benzoyl]-L-glutamic Acid Disodium Salt (Example 1 of U.S. Publication No. 2003/0216416)

Active ingredient:   4% (total solution) L-cysteine: 0.03% (total solution) A pharmaceutically acceptable excipient: water

The pH of the solution was adjusted to 8.5 using sodium hydroxide. The pH adjusted solution was protected from light. The solution was purged with nitrogen for twenty minutes and then sterile filtered. The formulation was dispensed into pre-washed, depyrogenated vials and then stoppered with a pre-washed, pre-sterilized teflon coated stopper. Caps were attached using a crimper. The sterile filtration and dispensing steps were conducted using a nitrogen isolator (5% v/v Oxygen). 

1. Pemetrexed diacid of the following formula

having a purity of at least about 99%.
 2. The pemetrexed diacid of claim 1, having a purity of at least about 99.7%.
 3. A process for preparing pemetrexed diacid having a purity of at least about 99% comprising: a) measuring the purity of the pemetrexed diacid in at least one batch of pemetrexed diacid; b) selecting a batch of pemetrexed diacid having a purity of at least about 98%; and c) preparing pemetrexed diacid having a purity of at least about 99% from the selected batch.
 4. The process of claim 3, wherein the measurement in step (a) is done by an HPLC method comprising: a) combining a pemetrexed diacid sample with a solution of NaH₂PO₄ to obtain a solution; b) injecting the solution into a 150×4.6 mm×3 μm Phenyl C18 or Octadecyl (C18) reversed phase silica gel column; c) eluting the sample from the column using a mixture of an aqueous solution of trifluoroacetic acid and acetonitrile; and d) measuring the pemetrexed diacid content in the sample with a UV detector.
 5. The process of claim 3, wherein the pemetrexed diacid having a purity of at least about 99% is prepared from the selected batch by a process comprising crystallizing pemetrexed diacid from a mixture comprising at least one solvent selected from the group consisting of amides, pyrrolidinones, and sulfoxides, at least one anti-solvent selected from the group consisting of alcohols, esters, ketones, water, halogenated hydrocarbons, aromatic hydrocarbons, and ethers, and optionally at least one co-solvent selected from the group consisting of nitriles; wherein the starting pemetrexed diacid has a purity of at least about 98%.
 6. The process of claim 5, wherein the solvent is selected from the group consisting of C₂₋₄ amides, C₄₋₆ pyrrolidinones, and C₂₋₄ sulfoxides.
 7. The process of claim 6, wherein the solvent is selected from the group consisting of dimethylformamide, dimethylacetamide, N-methyl pyrrolidinone, and dimethylsulfoxide.
 8. The process of claim 5, wherein the anti-solvent is selected from the group consisting of C₁₋₄ alcohols, C₃₋₆ esters, C₃₋₆ ketones, C₁₋₃ halogenated hydrocarbons, C₆₋₉ aromatic hydrocarbons, and C₄₋₈ ethers.
 9. The process of claim 8, wherein the anti-solvent is selected from the group consisting of methanol, ethanol, propanol, isopropanol, butanol, ethyl acetate, methyl acetate, isobutyl acetate, acetone, dichloromethane, toluene, and tetrahydrofuran.
 10. The process of claim 5, wherein the solvent is dimethylsulfoxide and the anti-solvent is ethanol.
 11. The process of claim 5, wherein the solvent is dimethylsulfoxide and the anti-solvent is a mixture of water and methanol.
 12. The process of claim 5, wherein the solvent is dimethylformamide and the anti-solvent is a mixture of water and methanol.
 13. The process of claim 5, wherein the co-solvent is a C₂₋₄ nitrile.
 14. The process of claim 13, wherein the co-solvent is acetonitrile.
 15. The process of claim 5, wherein the pemetrexed diacid having a purity of at least about 98%, the solvent, and optionally the co-solvent, are combined to form a combination, and the combination is heated to obtain a solution, prior to adding the anti-solvent.
 16. The process of claim 15, wherein the combination is heated to a temperature of about 25° C. to about 75° C. to obtain the solution.
 17. The process of claim 15, wherein the anti solvent is added to the solution at a temperature of about 30° C. to about 65° C.
 18. The process of claim 17, wherein the addition of the anti-solvent to the solution results in the formation of a suspension having a precipitate of pemetrexed diacid.
 19. The process of claim 18, wherein the suspension is maintained at a temperature of about 30° C. to about 65° C.
 20. The process of claim 5, further comprising recovering pemetrexed diacid that has a purity of at least about 99%.
 21. The process of claim 20, wherein the recovered pemetrexed diacid has a purity of at least about 99.7%.
 22. The process of claim 5, further comprising slurrying the crystallized pemetrexed diacid in the anti-solvent; and recovering the pemetrexed diacid from the slurry.
 23. The process of claim 22, wherein the pemetrexed diacid is slurried in the anti-solvent at a temperature of about 5° C. to about 65° C.
 24. The process of claim 22, wherein the anti-solvent is water, ethanol, methanol, or a mixture thereof.
 25. A process for preparing a pharmaceutically acceptable salt of pemetrexed diacid comprising preparing pemetrexed diacid having a purity of at least about 99% by the process of claim 3; and converting the pemetrexed diacid to the pharmaceutically acceptable salt of pemetrexed diacid.
 26. The process of claim 25, wherein the pharmaceutically acceptable salt of pemetrexed diacid is pemetrexed disodium.
 27. The process of claim 25, wherein the pharmaceutically acceptable salt of pemetrexed diacid has a purity of at least about 99%.
 28. A process for preparing a formulated pharmaceutically acceptable salt of pemetrexed diacid comprising preparing pemetrexed diacid having a purity of at least about 99% by the process of claim 3; and formulating the obtained pemetrexed diacid into a formulation comprising a pharmaceutically acceptable salt of pemetrexed diacid.
 29. The process of claim 28, wherein the formulated pharmaceutically acceptable salt of pemetrexed diacid is lyophilized pemetrexed disodium.
 30. The process of claim 28, wherein the pharmaceutically acceptable salt of pemetrexed diacid in the formulation has a purity of at least about 99%. 